Patients with sickle cell disease (SCD) have a manifest hypercoagulopathy. Multiple studies show that plasma from these patients exhibit: 1) increased thrombin generation; 2) decreased levels of natural anticoagulant proteins; and 3) abnormal activation of fibrinolysis. There also may be increased incidence of pulmonary embolism in SCD and histopathologic studies have suggested that large vessel narrowing with superimposed thrombosis is the most common cause of strokes in SCD. Several mechanisms have been proposed to underlie this hyper-coagulation including both inflammatory and hemolytic factors. Here we provide evidence of a novel mechanism of coagulation in SCD. Our previous work demonstrated that stress reticulocytes sRetics bind to blood monocytes via a fibronectin bridge to form a sRetic/monocyte aggregate. We now show that this binding directly induces monocyte tissue factor expression. We also found a strong correlation between stress reticulocyte count, the number of the sRetic/monocyte aggregates and tissue factor (TF) activity on plasma microparticles and markers of thrombin generation in these patients. Furthermore we show that erythropoietin (EPO), a cytokine involved in erythropoiesis, promotes the binding of the sRetic to the monocytes. Our data indicate EPO may associate with TF activity and thrombin generation in patients with SCD. Therefore, we propose to further explore the relationship between the sRetic-induced monocyte TF expression, EPO and coagulation activation in patients with SCD. In specific aim 1 we will examine the relationship between the sRetic/monocyte aggregate, monocyte TF expression/activity and established parameters of coagulation activation in SCD patients in the context of hemolysis and inflammation present in the disease. In aim 2, we will further define a role for EPO in coagulation activation and determine if EPO promotes coagulation in SCD by promoting sRetic/monocyte aggregate formation and subsequent TF expression. The studies will validate our in vitro work in patients and pave the way for further study of this novel mechanism of coagulation in SCD. PUBLIC HEALTH RELEVANCE: Hypercoagulation in sickle cell disease is likely a multi-factorial process involving multiple hemostatic parameters. However, these processes are still poorly understood. The more we understand about the etiology of this sickle-related coagulopathy, however, the more like we are to effectively target and treat thrombosis in this illness. In doing so, multiple sequelae of the illness, such as pulmonary embolism, deep vein thrombosis, liver and kidney infarct and pulmonary hypertension will be alleviated. We have discovered a potentially novel mechanism of coagulation activation in sickle cell disease, and this proposal will elucidate its relevance in the disorder.